Development and validation of a sensitive method for hydromorphone in human plasma by normal phase liquid chromatography–tandem mass spectrometry

Abstract

Liquid chromatography coupled with tandem mass spectrometry (LC–MS–MS) was developed for the quantitation of hydromorphone (HYD), an opiate analgesic, in human plasma. A simple liquid–liquid extraction was used to extract the analyte and its deuterated internal standard (d 3-HYD). Chromatographic separation of hydromorphone from its metabolite hydromorphone-3-glucuronide (H3G) was necessary because of the significant H3G fragmentation to HYD before Q1 of the mass spectrometer, which could result in false detection as HYD in the multiple reaction mode (MRM). This separation was achieved using a 50×2 mm, I.D. silica column (5 μm) and a mobile phase of acetonitrile–water–formic acid (80:20:1, v/v/v). The method was validated in the concentration range 0.05–10 ng ml −1 in plasma and met the acceptance criteria of industry guidelines for accuracy, precision, and stability.